Optimized method for the quantification of pyruvic acid in onions by microplate reader and confirmation by high resolution mass spectra

- Developed optimized microplate method for pyruvate analysis.
- KOH enhanced the stability of the colored hydrazone complex.
- Method is simple, rapid for the detection pyruvic acid in onions.
- First report on storage study of onion juice for pyruvate level.
- No significant difference in microplate and automated methods.

The present study describes the rapid microplate method to determine pyruvic acid content in different varieties of onions. Onion juice was treated with 2,4-dinitrophenylhydrazine to obtain hydrazone, which was further treated with potassium hydroxide to get stable colored complex. The stability of potassium complex was enhanced up to two hours and the structures of hydrazones were confirmed by LC-MS for the first time. The developed method was optimized by testing different bases, acids with varying concentrations of dinitrophenyl hydrazine to get stable color and results were comparable to developed method. Repeatability and precision showed <9% relative standard deviation. Moreover, sweet onion juice was stored for four weeks at different temperatures for the stability; the pyruvate remained stable at all temperatures except at 25 °C. Thus, the developed method has good potential to determine of pungency in large number of onions in a short time using minimal amount of reagents.